•A method to quantify ciprofloxacin in plasma by HPLC–UV.•Simple protein precipitation from serum with good recovery.•Validated method with acceptable reproducibility, precision, accuracy and stability.•Useful to quantify the concentration of ciprofloxacin in patients with Peripheral Arterial Disease.•Study will establish if an appropriate dosage regimen is being given to such patients. A rapid and sensitive HPLC–UV method for the determination of ciprofloxacin in human plasma is described. Protein precipitation with acetonitrile was used to separate the drug from plasma protein. An ACE® 5 C18 column (250mm×4.6mm, 5μm) with an isocratic mobile phase consisting of phosphate buffer (pH 2.7) and acetonitrile (77:23, v/v) was used for separation. The UV detector was set at 277nm. The method was validated in the linear range of 0.05–8μg/ml with acceptable inter- and intra-assay precision, accuracy and stability. The method is simple and rapid and can be used to quantify this widely used antibiotic in the plasma of patients suffering from Peripheral Arterial Disease.